In-situ growth of a spherical vinyl-functionalized covalent organic framework as stationary phase for capillary electrochromatography-mass spectrometry analysis.

Column technology is an important part in capillary electrochromatographic science. Developing novel stationary phase with high separation efficiency and high loading capacity is an essential work. In this work, a novel spherical vinyl-functionalized covalent-organic framework (COF-V) was synthesized at room temperature and firstly employed as stationary phase for CEC-MS analysis. The COF-V based CEC column was characterized by scanning electron microscopy, X-ray diffraction and Fourier transform infrared spectroscopy. The results proved the successful modification of COF-V. The COF-V based column possesses the advantages like strong electroosmotic flow, high separation efficiency and high loading capacity. The CEC column showed powerful separation selectivity to several kinds of compounds, and the highest column efficiency (theoretical plates, N) was over 1.4 × 105 plates·m-1 for methylbenzene. Besides, the COF-V modified column exhibited excellent repeatability and stability. The relative standard deviations (RSDs) of retention times for intra-day (n = 5), inter-day (n = 3) runs and column-to-column (n = 3) were all less than 2.1%. Hence, the COF-V modified column was successfully applied in CEC-MS for determination of antiepileptic drug, triazine herbicides and active ingredients in traditional Chinese medicine.

Electroosmotic extraction coupled to mass spectrometry analysis of metabolites in live cells.

Quantitative mass spectrometry analysis of metabolites at a single-cell level is critical to understanding the cell functionality and heterogeneity. To preserve cell viability after extraction, the extracted volume needs to be precisely controlled at a subpicoliter-to-picoliter level. Recently, we developed a volume-controlled, and highly sensitive approach for live cell analysis at a single-cell level by integrating electroosmotic extraction and nano-electrospray ionization mass spectrometry (nanoESI MS) analysis. Herein, we use outer epidermal cells of Allium cepa as a model system to present the details of our workflow, including detailed descriptions of the experimental setup for live cell analysis, preparation of the extraction nanopipette, establishment of calibration curves, and extraction and quantification of glucose in an individual onion cell. The capability of this procedure for quantitative live cell analysis has been demonstrated by accurate quantification of glucose in Allium cepa. In principle, our approach is applicable to identification and quantification of metabolites in live mammalian cells.

Quality evaluation of six bioactive constituents in goji berry based on capillary electrophoresis field amplified sample stacking.

Goji berry, fruits of the plant Lycium barbarum L., has long been used as traditional medicine and functional food in China. In this work, a simple and easy-operation on-line concentration capillary electrophoresis (CE) for detection flavonoids in goji berry was developed by coupling of field amplified sample stacking (FASS) with an electroosmotic (EOF) pump driving water removal process. Due to the EOF pump and electrokinetic injection showing different influence on the concentration, the analytes injection condition should be systemically studied. Thereafter, the verification of the analytes injection conditions was achieved using response surface experimental design. Under the optimum conditions, 86-271 folds sensitivity enhancement upon normal capillary zone electrophoresis (CZE, 50 mbar × 5 s) were achieved for six flavonoids, and the detection limits ranged from 0.35 to 1.82 ng/mL; the LOQ ranged from 1.20 to 6.01 ng/mL. Eventually, the proposed method was applied to detect flavonoids in 30 goji berry samples from different habitats of China; and the results indicated that the flavonoids were rich in the eluent of 30-60% methanol, which provided a reference for extraction of goji berry flavonoids.

Advanced portrayal of SMIL coating by allying CZE performance with in-capillary topographic and charge-related surface characterization.

A successive multiple ionic polymer layer (SMIL) coating composed of four layers improved the capillary electrophoretic separation of a recombinant major birch pollen allergen and closely related variants when poly(acrylamide-co-2-acrylamido-2-methyl-1-propansulfonate) (55% PAMAMPS) replaced dextran sulfate as terminal SMIL layer. 55% PAMAMPS decelerated the electroosmotic flow (EOF) due to its lower charge density. Atomic force microscopy (AFM) was used to investigate SMIL properties directly on the inner capillary surface and to relate them to EOF measurements and results of associated CZE separations of a mixture of model proteins and peptides that were performed in the same capillary. For the first time, AFM-based biosensing topography and recognition imaging mode (TREC) under liquid conditions was applied for a sequential characterization of the inner surface of a SMIL coated capillary after selected treatments including pristine SMIL, SMIL after contact with the model mixture, after alkaline rinsing, and the replenishment of the terminal polyelectrolyte layer. A cantilever with tip-tethered avidin was used to determine the charge homogeneity of the SMIL surface in the TREC mode. SMIL coated rectangular capillaries with 100 µm internal diameter assured accessibility of the inner surface for this cantilever type. Observed changes in CZE performance and EOF mobility during capillary treatment were also reflected by alterations in surface roughness and charge distribution of the SMIL coating. A renewal of the terminal SMIL layer restored the original surface properties of SMIL and the separation performance. The alliance of the novel TREC approach and CZE results allows for an improved understanding and a comprehensive insight in effects occurring on capillary coatings.

The electric field strength in orifice-like nanopores of ultrathin membranes.

Here we show that the electric field inside an ultrathin membrane is weaker than conventional theory would predict, and that the reduced field is predictive of measured electroosmotic flow rates. Our theoretical analysis shows that the electric field inside a charged nanopore is affected by end effects and dependent on the Dukhin number Du when the pore length-to-diameter aspect ratio λ is less than 80 for Du â‰ª 1 or 300 for Du â‰« 1. The electric field follows an unconventional scaling law; it no longer scales uniformly with the thickness of membrane, but with the local value of λ for each nanopore.

Anion separations with pressure-assisted capillary electrophoresis using a sequential injection analysis manifold and contactless conductivity detection.

It is demonstrated that a hydrodynamic flow superimposed on the mobility of analyte anions can be used for the optimization of analysis time in capillary zone electrophoresis. It was also possible to use the approach for counter-balancing the electroosmotic flow and this works as well as the use of surface modifiers. To avoid any band-broadening due to the bulk flow narrow capillaries of 10 µm internal diameter were employed. This was enabled by the use of capacitively coupled contactless conductivity detection, which does not suffer from the downscaling, and detection down to between 1 and 20 µM for a range of inorganic and small organic anions was found feasible. Precisely controlled hydrodynamic flow was generated with a sequential injection manifold based on a syringe pump. Sample injection was carried out with a new design relying on a simple piece of capillary tubing to achieve the appropriate back-pressure for the required split-injection procedure.

Gate manipulation of DNA capture into nanopores.

Understanding biophysics governing DNA capture into a nanopore and establishing a manipulation system for the capture process are essential for nanopore-based genome sequencing. In this work, the functionality of extended electric field and electroosmotic flow (EOF) during the capture stage and their dependence on gate voltage, U(G), are investigated. We demonstrate that while both the electric field and EOF within a cis chamber make long-distance contributions to DNA capture around the pore mouth, the former effect is always capturing, while the latter causes trapping or blocking of the molecule depending on the magnitude of the gate voltage, U(G): an anionic EOF induced by high U(G) is capable of doubling the DNA trapping speed and thus the absorption radius in the cis chamber, whereas a cationic EOF by low U(G) would substantially offset the trapping effort by the electric field and even totally block DNA entrance into the pore. Based on the analysis, a gate regulation is proposed with the objective of achieving a high DNA capture rate while maintaining a low error rate.

Highly efficient electroosmotic flow through functionalized carbon nanotube membranes.

Carbon nanotube membranes with inner diameter ranging from 1.5-7 nm were examined for enhanced electroosmotic flow. After functionalization via electrochemical diazonium grafting and carbodiimide coupling reaction, it was found that neutral caffeine molecules can be efficiently pumped via electroosmosis. An electroosmotic velocity as high as 0.16 cm s(-1) V(-1) has been observed. Power efficiencies were 25-110 fold improved compared to related nanoporous materials, which has important applications in chemical separations and compact medical devices. Nearly ideal electroosmotic flow was seen in the case where the mobile cation diameter nearly matched the inner diameter of the single-walled carbon nanotube resulting in a condition of using one ion is to pump one neutral molecule at equivalent concentrations.

A low-voltage nano-porous electroosmotic pump.

A low-voltage electroosmotic (EO) micropump based on an anodic aluminum oxide (AAO) nano-porous membrane with platinum electrodes coated on both sides has been designed, fabricated, tested, and analyzed. The maximum flow rate of 0.074 ml min(-1) V(-1) cm(-2) for a membrane with porosity of 0.65 was obtained. A theoretical model, considering the head loss along the entire EO micropump system and the finite electrical double layer (EDL) effect on the flow rate, is developed for the first time to analyze the performance of the EO micropump. The theoretical and experimental results are in good agreement. It is revealed that the major head loss could remarkably decrease the flow rate, which thus should be taken into account for the applications of the EO micropump in various Lab-on-a-chip (LOC) devices. However, the effect of the minor head loss on the flow rate is negligible. The resulting flow rate increases with increasing porosity of the porous membrane and kappaa, the ratio of the radius of the nanopore to the Debye length.

Comparison of aromatic and alkyl micelles for the electrokinetic determination of phthalates in virgin olive oil.

Two electrokinetic methods for the separation of phthalates are proposed. One uses 25 mM sodium borate and 50 mM sodium taurodeoxycholate adjusted to pH 2.8, and the other uses 15 mM ammonium tetraborate, 100 mM SDS, 0.25% w:v hydroxypropylmethyl cellulose (HPMC) and 5% v:v methanol (MeoH) adjusted to pH 9. The BGE containing SDS micelles as the pseudo-stationary phase provided better analytical figures of merit, particularly as regards LOD (0.4-1.4 mg/L) and precision (1.1-6.5%). Adding MeoH and HPMC to the BGE proved essential in order to obtain narrow and symmetric peaks. The proposed method was successfully used to determine phthalates in virgin olive oil. Recoveries from spiked samples ranged from 96 to 106%. The precision obtained in the analysis of real samples, as RSD, was better than 6.8%.

Microemulsion electrokinetic chromatography coupling with field amplified sample injection and electroosmotic flow suppressant for analysis of some quinolizidine alkaloids.

A new rapid and reproducible method using microemulsion electrokinetic chromatography (MEEKC) combining field amplified sample injection and electroosmotic flow suppressant for the analysis of five quinolizidine alkaloids is developed in this paper. For the separation of five quinolizidine alkaloids, a running buffer composed of 1.2% (v/v) 1-butanol, 0.6% (v/v) ethyl acetate and 98.2% (v/v) 1 mM Na(2)B(4)O(7)-2 mM NaH(2)PO(4) buffer solution containing 21 mM sodium cholate (SC) (pH 6.5) was developed. The resolution of the analytes was improved significantly by adding a divalent cation (e.g., Mg(2+)) to the running buffer as an electroosmotic flow modification. In order to analyze trace quinolizidine alkaloids in traditional Chinese herbal medicines, field amplified sample injection (FASI) was applied to increase the detection sensitivity. The detection limits (defined as S/N=3) for the analytes could be as low as 0.0001 microg/mL. This method was applied for the determination of quinolizidine alkaloids in real samples with simple extraction procedures, and the assay results were satisfactory.

Rapid determination of pK (a) values of 20 amino acids by CZE with UV and capacitively coupled contactless conductivity detections.

A rapid and universal capillary zone electrophoresis (CZE) method was developed to determine the dissociation constants (pK (a)) of the 20 standard proteogenic amino acids. Since some amino acids are poorly detected by UV, capacitively coupled contactless conductivity detection (C(4)D) was used as an additional detection mode. The C(4)D coupling proved to be very successful on a conventional CE-UV instrument, neither inducing supplementary analyses nor instrument modification. In order to reduce the analysis time for pK (a) determination, two strategies were applied: (i) a short-end injection to reduce the effective length, and (ii) a dynamic coating procedure to generate a large electroosmotic flow (EOF), even at pH values as low as 1.5. As a result, the analysis time per amino acid was less than 2 h, using 22 optimized buffers covering a pH range from 1.5 to 12.0 at a constant ionic strength of 50 mM. pK (a) values were calculated using an appropriate mathematical model describing the relationship between effective mobility and pH. The obtained pK (a) values were in accordance with the literature.

Analysis of nucleotides by pressure-assisted capillary electrophoresis-mass spectrometry using silanol mask technique.

A method for the determination of nucleotides based on pressure-assisted capillary electrophoresis-electrospray ionization mass spectrometry (PACE-MS) is described. To prevent multi-phosphorylated species from adsorbing onto the fused-silica capillary, silanol groups were masked with phosphate ions by preconditioning the capillary with the background electrolyte containing phosphate. During preconditioning, nebulizer gas was turned off to avoid contamination of MS detector with phosphate ions. To detect nucleotides using the CE positive mode at a pH 7.5, it was necessary to apply air pressure to the inlet capillary during electrophoresis to supplement the electroosmotic flow (EOF) toward the cathode. Moreover, we exchanged the running electrolyte every analysis using the buffer replenishment system to obtain the required reproducibility. Under the optimized conditions, 14 phosphorylated species such as nucleotides, nicotinamide-adenine dinucleotides and coenzyme A (CoA) compounds were well determined in less than 20 min. The relative standard deviations (n=6) of the method were better than 0.9% for migration times and between 1.7% and 8.1% for peak areas. The detection limits for these species were between 0.5 and 1.7 micromol/L with pressure injection of 50 mbar for 30 s (30 nL) at a signal-to-noise ratio of 3. This approach is robust and quantitative compared to the previous method, and its utility is demonstrated by the analysis of intracellular nucleotides and CoA compounds extracted from Escherichia coli wild type, pfkA and pfkB knockout mutants. The methodology was used to suggest that pfkA is the main functional enzyme.

Enhancement of skin permeation of high molecular compounds by a combination of microneedle pretreatment and iontophoresis.

A combination of microneedle pretreatment and iontophoresis was evaluated for the potential to increase skin permeation of drugs. Two model compounds with low and high molecular D(2)O and fluorescein isothiocyanate (FITC)-dextrans (FD-4, FD-10, FD-40, FD-70 and FD-2000; average molecular weight of 3.8, 10.1, 39.0, 71.2 and 200.0 kDa), respectively, were used and the effect of microneedle pretreatment and iontophoresis on their in vitro permeability was evaluated using excised hairless rat skin with a 2-chamber diffusion cell. Convective solvent flow through the skin was measured using a set of calibrated capillaries attached to the diffusion cell. The following results were obtained: (1) convective solvent flow (electroosmosis) during iontophoresis through microneedle-pretreated skin, 2.62+/-0.32 microL/cm(2)/h, was almost the same as through intact skin, 2.71+/-0.25 microL/cm(2)/h, and (2) the combination of microneedle pretreatment and subsequent iontophoresis significantly enhanced FD flux compared with microneedle pretreatment alone or iontophoresis alone, whereas no synergistic effect was found on the flux of D(2)O. These results suggest that the combination of iontophoresis with microneedle pretreatment may be a useful means to increase skin permeation of high molecular compounds.